Identification of a His-Asp-Cys Catalytic Triad Essential for Function of the Rho Inactivation Domain (RID) of Vibrio cholerae MARTX Toxin

Ahrens, Sebastian; Geissler, Brett; Satchell, K.J.F.

doi:10.1074/jbc.m112.396309

Cited by 32 publications

(46 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Alanine scanning mutagenesis has proven to be a useful tool to identify critical residues for other of MARTX effector domains 12,31,38 . Alignment of DUF5 amino acid sequences from 5 MARTX toxins, Plu2400, and PMT showed that there are only 16 residues (8.5%) that are 100% identical across all proteins.…”

Section: Resultsmentioning

confidence: 99%

“…The actin crosslinking domain (ACD) covalently links actin monomers via an isopeptide bond leading to actin cytoskeletal destruction 7–10 . The Rho GTPase inactivation domain (RID) disables the Rho regulatory pathway resulting in loss of active Rho and thereby to cytoskeleton depolymerization 11,12 . The ExoY domain is an adenylate cyclase 13 .…”

Section: Introductionmentioning

confidence: 99%

“…The C1 Pm subdomain from PMT is known to be a four helical bundled membrane localization domain (4HBM) 16 . The conserved C1 Vv subdomain from DUF Vv has also been demonstrated to localize to the eukaryotic plasma cell membrane, where it binds anionic lipids via a basic-hydrophobic motif 12,17 . Structural determination by nuclear magnetic resonance of the isolated C1 Pm and C1 Vv subdomains confirm both of these domains form a four helical bundle in solution 18,19 .…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Cytotoxicity of the V ibrio vulnificus MARTX toxin Effector DUF5 is linked to the C2A Subdomain

2014

View full text Add to dashboard Cite

The multifunctional-autoprocessing repeats-in-toxin (MARTX) toxins bacterial protein toxins that serve as delivery platforms for cytotoxic effector domains. The domain of unknown function in position 5 (DUF5) effector domain is present in at least six different species’ MARTX toxins and as a hypothetical protein in Photorhabdus spp. Its presence increases the potency of the Vibrio vulnificus MARTX toxin in mouse virulence studies, indicating DUF5 directly contributes to pathogenesis. In this work, DUF5 is shown to be cytotoxic when transiently expressed in HeLa cells. DUF5 localized to the plasma membrane dependent upon its C1 domain and the cells become rounded dependent upon its C2 domain. Both full-length DUF5 and the C2 domain caused growth inhibition when expressed in Saccharomyces cerevisiae. A structural model of DUF5 was generated based on the structure of Pasteurella multocida toxin facilitating localization of the cytotoxic activity to a 186 amino acid subdomain termed C2A. Within this subdomain, an alanine scanning mutagenesis revealed aspartate-3721 and arginine-3841 as residues critical for cytotoxicity. These residues were also essential for HeLa cell intoxication when purified DUF5 fused to anthrax toxin lethal factor was delivered cytosolically. Thermal shift experiments indicated that these conserved residues are important to maintain protein structure, rather than for catalysis. The Aeromonas hydrophila MARTX toxin DUF5Ah domain was also cytotoxic, while the weakly conserved C1-C2 domains from P. multocida toxin were not. Overall, this study is the first demonstration that DUF5 as found in MARTX toxins has cytotoxic activity that depends on conserved residues in the C2A subdomain.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Cytotoxicity of the V ibrio vulnificus MARTX toxin Effector DUF5 is linked to the C2A Subdomain

2014

View full text Add to dashboard Cite

show abstract

“…This effector is a member of the circular permuted thiol peptidase family, although the RID is predicted to have a distinct arrangement of its catalytic site compared to other proteins in this family (43, 44). After delivery to the cytosol by processing from CPD, the presence of a four helical bundle type membrane localization domain targets the RID to the plasma membrane (45, 46).…”

Section: Martx Toxins Of V Choleraementioning

confidence: 99%

“…As Rho is a master regulator of stress fiber assembly, cells treated with RID round due to lack of cytoskeletal assembly (47). Interestingly, although both ACD and RID independently affect polymerized actin, the domains are not synergistic, and thus, neither actin crosslinking nor Rho inactivation is accelerated by the presence of the other domain in the same toxin (26, 44). …”

Section: Martx Toxins Of V Choleraementioning

confidence: 99%

Multifunctional-autoprocessing repeats-in-toxin (MARTX) Toxins of Vibrios

Satchell

2015

Microbiol Spectr

112

View full text Add to dashboard Cite

Multifunctional-autoprocessing repeats-in-toxin (MARTX) toxins are a heterogeneous group of toxins found in a number of Vibrio species and other Gram-negative bacteria. The toxins are composed of conserved repeat regions and an autoprocessing protease domain that together function as a delivery platform for transfer of cytotoxic and cytopathic domains into target eukaryotic cell cytosol. Within the cells, the effectors can alter biological processes such as signaling or cytoskeletal structure, presumably to the benefit of the bacterium. Ten effector domains are found in the various Vibrio MARTX toxins, although any one toxin carries only two to five effector domains. The specific toxin variant expressed by a species can be modified by homologous recombination to acquire or lose effector domains, such that different strains within the same species can express distinct variants of the toxins. This review examines the conserved structural elements of the MARTX toxins and details the different toxin arrangements carried by Vibrio species and strains. The catalytic function of domains and how the toxins are linked to pathogenesis of human and animals is described.

show abstract